For cells, keeping quiet is a cinch

Researchers show how dormant cells turn genes off by pulling DNA into loops

Jan. 3, 2019

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By Sabrina Richards / Fred Hutch News Service

Cells use a protein called condensin (symbolized by a blue loop) to help them enter a paused state called quiescence.

Illustration by Kim Carney / Fred Hutch News Service

Stress can make you wish life had a pause button. Single-celled organisms like yeast actually have this option. Faced with a lack of food or other stressors, baker’s yeast can enter a “paused,” energy- and resource-conserving state called quiescence. In this paused state, in which DNA becomes more compact and most genes are shut off, yeast can live weeks or months without nutrients.

A key step in putting normal life on hold is cinching DNA into regular loops, according to new work by researchers at Fred Hutchinson Cancer Research Center, the University of Washington and the University of Massachusetts. In a study published today in the journal Molecular Cell, they identified the protein that yeast cells use to tighten DNA and shut off most genes.

The findings resolve a chicken-and-egg debate of the quiescence field: Does DNA become compact because most genes are shut off, or is DNA compaction a strategy to help turn genes off?

According to the new study, DNA compaction is a strategy, not a side effect. Genes get turned off because DNA compacts.

Some of our own cells, like the stem cells that feed our tissues, also temporarily enter this restful state under normal conditions. And evidence suggests that cancer cells may be able to resist anti-cancer drugs by becoming similarly dormant. A better understanding of this paused state could help scientists discover ways to awaken dormant tumor cells or sensitize them to anti-cancer drugs.

Dr. Toshio Tsukiyama

Fred Hutch file

The next DNA dimension

If the DNA from a single human cell were unraveled, it would extend about six feet. The key to fitting this neatly into our cells is packaging. Packaged DNA is called chromatin. The most basic layer of chromatin, in which DNA loops around a cartwheel-shaped protein, is known as beads on a string. A lot is known about how changes to these beads can turn genes on or off.

“It’s been known for a really long time that chromatin structure plays a very important role in basically everything that happens to DNA,” said Dr. Toshio Tsukiyama, a Fred Hutch genome scientist and senior author of the paper. Whether genes get turned on, when DNA is replicated — these and other fundamental processes are influenced by chromatin.

But this simple winding isn’t enough to compress DNA enough to fit inside the nucleus, and it undergoes further looping and folding. Less is known about how this 3D chromatin structure may influence cellular processes. Work by others has suggested that the folding doesn’t occur at random, and that it may create specific sections, or domains, of chromatin. These domains may act to enhance interactions between certain segments of DNA while reducing interactions for others, but researchers lacked strong evidence for direct roles of chromatin domains.

Driven by technology

“This is the first report to identify a factor important for chromatin compaction in quiescent cells,” and to suggest that chromatin compaction functions to keep genes turned off, rather than the other way around, Tsukiyama said.

A breakthrough technology, called Micro-C, made the current findings possible. Micro-C was developed by two members of the research team, Dr. Oliver Rando and graduate student Tsung-Han Hsieh at the University of Massachusetts. A previous approach gave researchers a much lower-resolution map of chromatin interactions. In map terms, it could show whether two genes share a large, city-sized chromatin domain. That resolution is much too low for researchers to see whether the domains they charted had any relationship to whether genes are off or on. Micro-C, on the other hand, can tell them if two genes share the same neighborhood.

Using Micro-C, Rando and Hsieh were the first to describe small 3D chromatin domains in baker’s yeast. In the current study, the collaborators used the next generation of Micro-C, Micro-C XL, to map out both smaller and larger domains in quiescent yeast cells.

Compaction comes first

Armed with Micro-C XL, Tsukiyama and Dr. Sarah Swygert, a postdoctoral fellow in his lab, set about looking at the 3D structure of chromatin in quiescent yeast cells compared to non-quiescent, quickly dividing yeast cells.

Their findings suggested that yeast cells create chromatin loops — domains with defined boundaries — that reduce interactions between sections of DNA in different regions. In quiescent cells, these boundaries appeared much firmer, suggesting that the loops are more isolated in dormant yeast.

By comparing Micro-C XL data to which genes are turned or or off during dormancy, Tsukiyama and Swygert saw that chromatin domain boundaries tend to fall in between a gene that is on during quiescence and a gene that is turned off. This suggested that the boundaries might play a role in gene activity during quiescence.

Illustration by Kim Carney / Fred Hutch News Service

Looking for the method cells use to set up the loops, the scientists turned to a handful of molecules known to be involved in 3D chromatin structure. One, called condensin for its role in condensing chromatin, showed a dramatic difference in where it bound to DNA in active cells compared to paused, quiescent cells. In dividing cells, condensin is concentrated in just two spots. But after yeast cells press their internal pause buttons, it becomes strewn across the DNA, binding at the boundaries of nearly every loop that the team had identified using Micro-XL.

And like those borders, condensin’s binding pattern also corresponded with the few genes that are turned on in paused cells.

The wholesale migration of condensin “was a big surprise for us,” Tsukiyama said. This, combined with the fact that its new location corresponded with the few active quiescence genes, “suggests that condensin is doing something to prevent interactions between those domains.”

To find out exactly what condensin is doing in dormant cells, the team developed a method to remove it as cells entered their quiet state. This caused big changes in chromatin compaction and which genes were off.

Without condensin, chromatin in quiescent cells was looser, and many of the genes within the loops did not shut off as they normally do in quiescence, suggesting that condensin-dependent chromatin compaction helps turn genes off.

Because quiescence is such a universal process, the researchers suspected that condensin might play a similar role in other living things, like humans. When they looked at human cells, they saw that chromatin was much looser in quiescent cells in which they’d blocked condensin’s activity, compared to quiescent cells with active condensin.

Quiescent cells need to conserve energy and resources. Tsukiyama and Swygert’s findings suggest that condensin-created loops help them do this by blocking activation of genes that are unneeded or even harmful during this dormant state.

Future steps

Tsukiyama is already looking to expand the work to other types of cells, including mammalian cells, to see how similar chromatin compaction processes are between different species. Though it’s a universal process, quiescence has been adapted by different species and different cell types to meet specific needs. Some insights, such as those concerning dormant cancer cells, could have big implications for human health.

Will human cells have novel chromatin domains, or different domain boundaries? Will condensin play a role, and if so, how big?

“It’s all on the table,” Tsukiyama said.

Sabrina Richards, a staff writer at Fred Hutchinson Cancer Research Center, has written about scientific research and the environment for The Scientist and OnEarth Magazine. She has a Ph.D. in immunology from the University of Washington, an M.A. in journalism and an advanced certificate from the Science, Health and Environmental Reporting Program at New York University. Reach her at srichar2@fredhutch.org.

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